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A preparation method of hemagglutination inhibitory antigen of Avian bacillus paragallinarum and application of the antigen

A hemagglutination inhibition, chicken and poultry technology, applied in the field of preparation of hemagglutination inhibition antigen, can solve the problems of low antigen titer, loss of hemagglutination activity, economic loss, etc., and achieve good precipitation and aggregation, excellent hemagglutination activity, The effect of high blood coagulation titer

Active Publication Date: 2020-09-29
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After the disease occurs, the egg production of laying hens will decrease, the growth and development of rearing chickens will be hindered, the elimination rate will increase, and the quality of broiler chickens will decrease, causing great economic losses.
[0003] In the existing serological detection method of Avian bacillus paragallinarum, KSCN / Nacl is often used as the antigen to treat the bacteria, and the antigen titer is low under the same amount of bacteria, and the antigen is unstable, and there is a phenomenon of losing hemagglutination activity. Very short shelf life, uneconomical

Method used

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  • A preparation method of hemagglutination inhibitory antigen of Avian bacillus paragallinarum and application of the antigen
  • A preparation method of hemagglutination inhibitory antigen of Avian bacillus paragallinarum and application of the antigen
  • A preparation method of hemagglutination inhibitory antigen of Avian bacillus paragallinarum and application of the antigen

Examples

Experimental program
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Effect test

Embodiment 1

[0050] Example 1 Preparation and Preservation of the Hemagglutination Inhibiting Antigen of Avibacterium paragallinarum

[0051] 1. Take Avibacterium paragallinarum (provided by the Animal Husbandry and Veterinary Laboratory of Beijing Academy of Agriculture and Forestry Sciences), mark it on a TSA plate, and place it in a candle jar at 37°C for 16h-28h, until the colonies grow out. Pick colonies from the purely cultured plate, inoculate 100mL TSB medium, incubate at 37°C for 10h-16h, harvest the culture when OD600≥0.9, centrifuge the culture at 8000×g for 10min, wash the precipitate with PBS for 3 times, Then suspend with 20mL-30mL PBS solution to obtain PBS suspension of Avibacterium paragallinarum.

[0052] 2. Take Staphylococcus suis (provided by the Institute of Animal Husbandry and Veterinary Medicine, Beijing Academy of Agriculture and Forestry Sciences), streak culture on ordinary broth agar medium, inoculate into LB liquid medium for overnight culture after the coloni...

Embodiment 2

[0056] Example 2 Preparation of 10% Aldehydelated Red Blood Cells

[0057] 1) Preparation of 10% fresh erythrocytes. The erythrocytes collected from healthy non-immune roosters were mixed with Alfred’s solution at a volume ratio of 1:1. After centrifugation at 500×g, the precipitate was washed with normal saline for 5 times. 10 times the volume of red blood cells, each centrifugation × 500g, centrifuged for 10 minutes, and the precipitated red blood cells were made into a 10% red blood cell suspension with a volume ratio of phosphate buffer (PB solution).

[0058] 2) Prepare glutaraldehyde solution (50%) according to the ratio of 5% of the above red blood cell suspension volume. First, dilute the glutaraldehyde stock solution from 50% to a final concentration of 5% glutaraldehyde with PBS, and place on a magnetic Put it on the stirrer, add it into the red blood cell suspension while stirring (if the volume of the red blood cell suspension is 100 ml, then take 5 ml of 50% gluta...

Embodiment 3

[0064] Embodiment 3 Hemagglutination (HA) test and hemagglutination inhibition (HI) test of Avian bacilli paragallinarum

[0065] 1. Hemagglutination (HA) test

[0066] The operation is best carried out at room temperature within the range of 20°C to 25°C. If the room temperature is not within this range, it can also be placed in a 37°C incubator. Take care to cover with a lid or parafilm to prevent the liquid from evaporating.

[0067] 1) HA test operation method: use a 96-well V-shaped microplate for the test, and the total reaction volume is 75 μl. According to Table 1, use bovine serum albumin PBS (abbreviated as PBSS, its configuration method is: take 0.1g bovine serum albumin V, dissolve it in 100ml PBS, filter and sterilize, put 2~8 ℃ for preservation.) Example 1 The antigen prepared in the method was diluted 5-fold first, then serially diluted 2-fold, then added 1% aldehylated red blood cells, mixed gently, and left at room temperature (15-25°C) for 40-60 minutes or ...

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Abstract

The invention provides a hemagglutination inhibition antigen of avibacterium paragallinarum, and a preparation method and application of the antigen. The antigen is prepared by adding staphylococcus hyicus filtrate into the avibacterium paragallinarum and treating. The hemagglutination inhibition antigen of the avibacterium paragallinarum can be applied to detection of an avibacterium paragallinarum antibody, and specifically can be used for preparing a reagent and a kit for a hemagglutination test or a hemagglutination inhibition test, and other immunological detection methods and reagents. The prepared hemagglutination inhibition antigen of the avibacterium paragallinarum is stable and has high hemagglutination titer; when the hemagglutination inhibition antigen is used for the hemagglutination test or the hemagglutination inhibition test, a result is more stable and easy to judge and the operability is strong.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a preparation method of a hemagglutination-inhibiting antigen of Avian paragallinarum and the application of the antigen. Background technique [0002] Avibacterium paragallinarum (A. paragallinarum) is the pathogenic bacterium of chicken infectious rhinitis, which can cause acute upper respiratory tract infectious diseases in chickens. The acute upper respiratory infectious disease of chicken all has generation and popular in many places in the world, and my country just has the case appearance of many suspected primary diseases from 1980, at first by Feng Wenda isolates bacterium in Beijing in 1987. After the disease occurs, it can cause the egg production of laying hens to decline, the growth and development of rearing chickens to be hindered and the elimination rate to increase, and the quality of broiler chickens to decline, causing great economic losses. [0003] I...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12N1/06C12N1/04G01N33/569C12R1/01
CPCC12N1/04C12N1/06C12N1/20G01N33/56911
Inventor 孙惠玲陈小玲程晶徐福洲
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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